Project Summary/Abstract Obesity is a primary risk factor for cardiovascular disease and metabolic disorders such as diabetes mellitus. Obesity and its metabolic consequences continue to be among the most important biomedical challenges in the U.S. and worldwide today. Nearly one-third of adults (33.8%) and 17% (or 12.5 million) of children and adolescents are obese in the U.S.. In the absence of improved therapies, further increases are expected in the number of serious conditions closely associated with obesity. All anti-obesity medications currently approved by the FDA act to repress energy intake, either by suppressing appetite or by inhibiting intestinal fat absorption. However, due to side effects including depression, oily bowel movements and steatorrhea, there is an urgent need for alternative approaches. This study is significant because it may elucidate new drug to counteract obesity by the completely opposite method, namely, via increasing energy expenditure. Two types of fat tissue exist in mammals. Brown adipose tissue (BAT) is a specialized adipose tissue that dissipate energy for heat generation, whereas white adipose tissue (WAT) functions as storage of excess energy. Studies suggest that loss of BAT is linked to decreased energy expenditure and obesity in humans; thus ?increasing? energy expenditure through regeneration of BAT could be effective to counteract obesity. Certain physiological cues, such as chronic cold exposure, convert WAT into mitochondria-rich, energy consuming BAT-like adipocyte. This ?browned? adipocyte is referred to as a ?beige adipocyte? and recent studies including ours indicate that adult human BAT is mostly composed of beige adipocytes. The objective of the proposed research is to determine all subtypes of cells that give rise to beige adipocytes. In this project, I will focus on cell types that are specific to subcutaneous WAT. It has been known that subcutaneous WAT can easily be ?browned? by external cues, whereas, this conversion is much less observed in a visceral WAT. I hypothesize that there are discreet cell types in subcutaneous WAT that are predetermined to differentiate to beige adipocytes. To test this hypothesis, I will employ a novel methodology (single cell RNA sequencing) and determine the number and characteristics of cell types in subcutaneous and visceral white adipose tissue. I then identify genetic biomarker for the depot-specific populations. These biomarkers will be used for the prospective isolation of stem cell of beige adipocytes which contribute to the development of novel cell based therapeutics or pharmacological strategies to counteract obesity in humans.